Quantitative X-Ray Elemental Mapping of Dynamic Physiologic Events in Skeletal Muscle

Abstract

A striking event in animal biology, both fundamental and universal, is the process of coupling excitation to contraction (ECC) in muscle. The first step in that process, the excitatory stimulus, is electrical and is followed by the release of free calcium from known intracellular stores called the “junctional SR” (JSR, Sommer, 1968). The freed calcium unlocks a cocked state of an actomyosin interaction at rest, causing the muscle to contract. Neither the mechanism by which the electrical signal is translated into calcium release, nor its precise time course are known. Whereas our initial aim has been to define the accurate time of that calcium release in single, intact frog skeletal muscle fibers (R. temporaria), our overall goal is the description of the quantitative changes in elemental microtopochemistry during the entire time-course of excitation-contraction coupling, in both skeletal and cardiac muscle. By measuring calcium directly, we are in a position to avoid the complexities introduced by as yet poorly-understood interactions between calcium and a number of indicator substances (Blinks, 1978; Baylor, 1988) that result in their emitting measurable calcium-dependent signals. Given the present model, we also have the unique opportunity to explore the limits, per se, of electron probe x-ray microanalysis (EPXMA) as a legitimate means for the study of dynamic physiologic events dominated by spatial displacements of elements. Single intact cells, excellent time resolution and optimal cryotechniques are imperative for such a purpose (Nassar, 1986; Sommer, 1988). Skeletal muscle fibers were used because they have many very large JSRs containing very high concentrations of calcium (Somlyo, 1980).